LOCUS Exported 4375 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Bacterial NM repression YFP reporter: protospacer A. ACCESSION . VERSION . KEYWORDS SK-YFP-NM-A SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4375) AUTHORS Esvelt KM, Mali P, Braff JL, Moosburner M, Yaung SJ, Church GM TITLE Orthogonal Cas9 proteins for RNA-guided gene regulation and editing. JOURNAL Nat Methods. 2013 Sep 29. doi: 10.1038/nmeth.2681. PUBMED 24076762 REFERENCE 2 (bases 1 to 4375) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4375 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(159..187) /label=GFP-R /note="GFP, reverse primer. Does NOT anneal to EGFP" primer_bind 782..802 /label=GFP-F /note="GFP, forward primer. Does NOT anneal to EGFP" terminator 904..990 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" CDS complement(1525..2475) /codon_start=1 /gene="rep101" /product="RepA protein needed for replication with the pSC101 origin" /label=Rep101 /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI" rep_origin 2523..2745 /label=pSC101 ori /note="low-copy replication origin that requires the Rep101 protein" terminator 3237..3331 /label=lambda t0 terminator /note="transcription terminator from phage lambda" CDS complement(3362..4156) /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(3473..3492) /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind 4083..4102 /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4335..4353 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" ORIGIN 1 gatccgataa atatctaaca ccgtgcgtgt tgactatttt acctctggcg gtgataatgg 61 ttgcatgtac tagaatcttt ttcaacaagc ttgagatggt actttaactt taagaaggag 121 atatacatat gcgtaaagga gaagaacttt tcactggagt tgtcccaatt cttgttgaat 181 tagatggtga tgttaatggg cacaaatttt ctgtcagtgg agagggtgaa ggtgatgcaa 241 catacggaaa acttaccctt aaatttattt gcactactgg aaaactacct gttccatggc 301 caacacttgt cactactttc ggttatggtc taaaatgctt tgctagatac ccagatcata 361 tgaaacggca tgactttttc aagagtgcca tgcccgaagg ttatgtacag gaaagaacta 421 tatttttcaa agatgacggg aactacaaga cacgtgctga agtcaagttt gaaggtgata 481 cccttgttaa tagaatcgag ttaaaaggta ttgattttaa agaagatgga aacattcttg 541 gacacaaatt ggaatacaac tataactcac acaatgtata catcatggca gacaaacaaa 601 agaatggaat caaagttaac ttcaaaatta gacacaacat tgaagatgga agcgttcaac 661 tagcagacca ttatcaacaa aatactccaa ttggcgatgg ccctgtcctt ttaccagaca 721 accattacct gtcctatcaa tctgcccttt cgaaagatcc caacgaaaag agagaccaca 781 tggtccttct tgagtttgta acagctgctg ggattacaca tggcatggat gaactataca 841 aataagctta accgaagcgt ttgatagttg atatcctttg cctgcggccg caactagagg 901 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg 961 tcggtgaacg ctctcctgag taggacaaat ccgccgccct agacctaggg tacgggtttt 1021 gctgcccgca aacgggctgt tctggtgttg ctagtttgtt atcagaatcg cagatccggc 1081 ttcaggtttg ccggctgaaa gcgctatttc ttccagaatt gccatgattt tttccccacg 1141 ggaggcgtca ctggctcccg tgttgtcggc agctttgatt cgataagcag catcgcctgt 1201 ttcaggctgt ctatgtgtga ctgttgagct gtaacaagtt gtctcaggtg ttcaatttca 1261 tgttctagtt gctttgtttt actggtttca cctgttctat taggtgttac atgctgttca 1321 tctgttacat tgtcgatctg ttcatggtga acagctttaa atgcaccaaa aactcgtaaa 1381 agctctgatg tatctatctt ttttacaccg ttttcatctg tgcatatgga cagttttccc 1441 tttgatatct aacggtgaac agttgttcta cttttgtttg ttagtcttga tgcttcactg 1501 atagatacaa gagccataag aacctcagat ccttccgtat ttagccagta tgttctctag 1561 tgtggttcgt tgtttttgcg tgagccatga gaacgaacca ttgagatcat gcttactttg 1621 catgtcactc aaaaattttg cctcaaaact ggtgagctga atttttgcag ttaaagcatc 1681 gtgtagtgtt tttcttagtc cgttacgtag gtaggaatct gatgtaatgg ttgttggtat 1741 tttgtcacca ttcattttta tctggttgtt ctcaagttcg gttacgagat ccatttgtct 1801 atctagttca acttggaaaa tcaacgtatc agtcgggcgg cctcgcttat caaccaccaa 1861 tttcatattg ctgtaagtgt ttaaatcttt acttattggt ttcaaaaccc attggttaag 1921 ccttttaaac tcatggtagt tattttcaag cattaacatg aacttaaatt catcaaggct 1981 aatctctata tttgccttgt gagttttctt ttgtgttagt tcttttaata accactcata 2041 aatcctcata gagtatttgt tttcaaaaga cttaacatgt tccagattat attttatgaa 2101 tttttttaac tggaaaagat aaggcaatat ctcttcacta aaaactaatt ctaatttttc 2161 gcttgagaac ttggcatagt ttgtccactg gaaaatctca aagcctttaa ccaaaggatt 2221 cctgatttcc acagttctcg tcatcagctc tctggttgct ttagctaata caccataagc 2281 attttcccta ctgatgttca tcatctgagc gtattggtta taagtgaacg ataccgtccg 2341 ttctttcctt gtagggtttt caatcgtggg gttgagtagt gccacacagc ataaaattag 2401 cttggtttca tgctccgtta agtcatagcg actaatcgct agttcatttg ctttgaaaac 2461 aactaattca gacatacatc tcaattggtc taggtgattt taatcactat accaattgag 2521 atgggctagt caatgataat tactagtcct tttcctttga gttgtgggta tctgtaaatt 2581 ctgctagacc tttgctggaa aacttgtaaa ttctgctaga ccctctgtaa attccgctag 2641 acctttgtgt gttttttttg tttatattca agtggttata atttatagaa taaagaaaga 2701 ataaaaaaag ataaaaagaa tagatcccag ccctgtgtat aactcactac tttagtcagt 2761 tccgcagtat tacaaaagga tgtcgcaaac gctgtttgct cctctacaaa acagacctta 2821 aaaccctaaa ggcttaagta gcaccctcgc aagctcgggc aaatcgctga atattccttt 2881 tgtctccgac catcaggcac ctgagtcgct gtctttttcg tgacattcag ttcgctgcgc 2941 tcacggctct ggcagtgaat gggggtaaat ggcactacag gcgcctttta tggattcatg 3001 caaggaaact acccataata caagaaaagc ccgtcacggg cttctcaggg cgttttatgg 3061 cgggtctgct atgtggtgct atctgacttt ttgctgttca gcagttcctg ccctctgatt 3121 ttccagtctg accacttcgg attatcccgt gacaggtcat tcagactggc taatgcaccc 3181 agtaaggcag cggtatcatc aacaggctta cccgtcttac tgtccctagt gcttggattc 3241 tcaccaataa aaaacgcccg gcggcaaccg agcgttctga acaaatccag atggagttct 3301 gaggtcatta ctggatctat caacaggagt ccaagcgagc tctcgaaccc cagagtcccg 3361 ctcagaagaa ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga 3421 taccgtaaag cacgaggaag cggtcagccc attcgccgcc aagctcttca gcaatatcac 3481 gggtagccaa cgctatgtcc tgatagcggt ccgccacacc cagccggcca cagtcgatga 3541 atccagaaaa gcggccattt tccaccatga tattcggcaa gcaggcatcg ccatgggtca 3601 cgacgagatc ctcgccgtcg ggcatgcgcg ccttgagcct ggcgaacagt tcggctggcg 3661 cgagcccctg atgctcttcg tccagatcat cctgatcgac aagaccggct tccatccgag 3721 tacgtgctcg ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa 3781 gcgtatgcag ccgccgcatt gcatcagcca tgatggatac tttctcggca ggagcaaggt 3841 gagatgacag gagatcctgc cccggcactt cgcccaatag cagccagtcc cttcccgctt 3901 cagtgacaac gtcgagcaca gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc 3961 gcgctgcctc gtcctgcagt tcattcaggg caccggacag gtcggtcttg acaaaaagaa 4021 ccgggcgccc ctgcgctgac agccggaaca cggcggcatc agagcagccg attgtctgtt 4081 gtgcccagtc atagccgaat agcctctcca cccaagcggc cggagaacct gcgtgcaatc 4141 catcttgttc aatcatgcga aacgatcctc atcctgtctc ttgatcagat cttgatcccc 4201 tgcgccatca gatccttggc ggcaagaaag ccatccagtt tactttgcag ggcttcccaa 4261 ccttaccaga gggcgcccca gctggcaatt ccgacgtcta agaaaccatt attatcatga 4321 cattaaccta taaaaatagg cgtatcacga ggccctttcg tcttcacctc gaggg //